MRP1 not MDR1 gene expression is the predominant mechanism of acquired multidrug resistance in two prostate carcinoma cell lines

Multidrug resistant prostate cancer cell lines DU 0.03 and PC 0.03 were established from the parental prostate cancer cell lines DU145 and PC-3 respectively by stepwise selection in doxorubicin (DOX) from 0.001 to 0.03 μg/ml. As cells adapted to each concentration of DOX. the drug concentration was increased by 0.001 μg/ml. The chemosensitivity of each line was determined by growth inhibition assay. The DU 0.03 and PC 0.03 lines exhibit a 5–10-fold and 1.3–2.8-fold increase in resistance to anthracyclines, vinblastine (VLB) and mitozantrone (Mito), respectively. Verapamil (5 μM) partially reversed the resistance to the anthracycline and completely reversed the resistance to VLB and Mito. Drug kinetic studies measured by intracellular accumulation of 3H-daunorubicin demonstrated a 3 fold decrease in the level of intracellular 3H-daunorubicin in the PC 0.03 and DU 0.03 resistant lines compared with their respective parental line. This effect was partially reversed by 5 μM verapamil. The expression of MDR1 and MRP genes was analysed by Northern blotting and RT-PCR. P-glycoprotein (Pgp) and MRP protein were tested by immunocytochemistry staining using the monoclonal antibodies J-SB1. C219 and MRK16 (Pgp) and MRPm6 and MRPr1 (MRP). Neither Northern blot analysis nor the more sensitive RT-PCR demonstrated detectable MDR1 transcripts in any of the prostate cancer cell lines and the three Pgp monoclonal antibodies failed to reveal expression of Pgp.